Aim To investigate the effect of miR-224-5p on the expression of proprotein convertase subtilisin/kexin type 9 (PCSK9) and the lipid uptake of HepG2 cells. Methods The position, conservativeness and seed sequence of miR-224-5p gene were analyzed by bioinformatics method. The binding sites and binding free energies of miR-224-5p to PCSK9 were analyzed in databases such as Targetscan, miRanda, miRDB and RNAhybrid. Direct targeted binding of miR-224-5p to PCSK9 mRNA 3′UTR was verified by double luciferase reporter gene. Western blot was used to detect the effects of miR-224-5p mimic and miR-224-5p inhibitor on PCSK9 and low density lipoprotein receptor (LDLR) proteins. LDLR on the cell membrane was directly observed with cellular immunofluorescence. Oil red O staining and DiI-LDL were respectively used to observe the effects of miR-224-5p on lipid droplet content and lipid uptake in HepG2 cells. Results Bioinformatics analysis revealed that the human miR-224-5p gene was located in Xq28 and highly conserved among different species. MiR-224-5p and PCSK9 mRNA 3′UTR had the basis of targeted binding, and the binding free energy was low. The double luciferase reporter gene assay showed that miR-224-5p mimic could inhibit the luciferase activity of PCSK9-WT 3′UTR, but not PCSK9-Mut 3′UTR, suggesting that the PCSK9 mRNA 3′UTR was the target of miR-224-5p. Further experiments showed that the miR-224-5p mimic could significantly inhibit the expression of PCSK9 protein and increase the content of LDLR protein. The expression of PCSK9 increased and the level of LDLR decreased after the down-regulation of miR-224-5p. In addition, oil red O staining showed that lipid droplets in HepG2 cells decreased significantly in the miR-224-5p mimic group, while lipid droplets in HepG2 cells increased significantly in the miR-224-5p inhibitor group. High expression of miR-224-5p significantly promoted LDLC uptake by HepG2 cells. Conclusion miR-224-5p targeting PCSK9 mRNA 3′UTR inhibits the expression of PCSK9, thereby reducing the content of lipid droplets in HepG2 cells and increasing the uptake of lipid by HepG2 cells.